PROJECT SUMMARY A growing body of evidence supports the idea that brown/beige adipose tissue (BAT) contributes to basal metabolism in adult humans, and that diminished BAT activity can lead to obesity. Sympathetic nerve stimulation increases BAT activity in rodents. Reports that signaling from the stellate ganglion (SG) modulates thermogenesis in humans, raises the possibility that minimally-invasive approaches to stimulate sympathetic projections from the SG to supraclavicular BAT (scBAT) have potential as an anti-obesity therapy. The neuroanatomy of the SG is complex, because sympathetic and sensory neuronal axons leave the ganglion through 6-8 different exit points to reach a wide variety of targets in the periphery. For neuromodulation of SG projections to BAT to have clinical applications, it is essential that stimulation protocols avoid projections from the SG to the heart and upper extremities. In theory, this can be achieved by physically focusing the electrical stimulus to SG neurons as they exit the SG or enter the scBAT depot. Alternatively, genetic approaches could be used restrict pharmacological manipulations to the subpopulation of SG neurons that innervates BAT. These complementary studies in mouse models and human tissue samples will serve as the basis for deciding which of these strategies are viable therapeutic options. The proposed studies are designed to fill critical information gaps and to develop tools needed for comprehensive mapping of neural circuits regulating BAT and to explore the potential use of BAT neuromodulation as an anti- obesity therapy. Studies in Aim 1 will use combinations of transgenic mouse models and fluorescent neuronal tracers to determine whether there is any physical overlap between BAT and forelimb-projecting soma within the SG or their exit points out of the SG. In parallel, we will perform the first mapping studies of the projections from the SG to scBAT in human autopsy specimens. A major obstacle to studies to modulate or record neural activity in the SG is that this ganglion contains many different types of afferent and efferent neurons that regulate a wide range of physiological functions. Studies in Aim 2 will define molecular markers for distinct subpopulations of SG neurons that project to BAT and heart. Then we will determine whether any of these markers are conserved in human surgical samples. Finally, studies in Aim 3 will establish systems to evaluate the impact of neuromodulation on the organization of sympathetic fibers in conjunction with well-established assays to measure BAT oxidative capacity and activity. To aid these efforts, we will develop techniques to image sympathetic projections in an intact BAT depot. In addition, we will validate key in vivo assays needed to assess the impacts of neuromodulation on BAT function that can be readily translated to humans. In addition to impacts on the BAT field, these studies will also provide a strong foundation for future efforts to understand how existing SG neuromodulation therapies have beneficial effects on a wide range of conditions that are refractory to other treatments, including ventricular tachycardia, chronic regional pain syndrome and post-traumatic stress disorder.